A panel of recombinant monoclonal antibodies against zebrafish neural receptors and secreted proteins suitable for wholemount immunostaining

Nicole Staudt; Nicole Müller-Sienerth; Alla Fane-Dremucheva; Shahnaz P Yusaf; David Millrine; Gavin J Wright

doi:10.1016/j.bbrc.2014.11.123

A panel of recombinant monoclonal antibodies against zebrafish neural receptors and secreted proteins suitable for wholemount immunostaining

Nicole Staudt, Nicole Müller-Sienerth, Alla Fane-Dremucheva, Shahnaz P Yusaf, David Millrine, Gavin J Wright

Research output: Contribution to journal › Article › peer-review

Abstract

Cell surface receptors and secreted proteins play important roles in neural recognition processes, but because their site of action can be a long distance from neuron cell bodies, antibodies that label these proteins are valuable to understand their function. The zebrafish embryo is a popular vertebrate model for neurobiology, but suffers from a paucity of validated antibody reagents. Here, we use the entire ectodomain of neural zebrafish cell surface or secreted proteins expressed in mammalian cells to select monoclonal antibodies to ten different antigens. The antibodies were characterised by Western blotting and the sensitivity of their epitopes to formalin fixation was determined. The rearranged antigen binding regions of the antibodies were amplified and cloned which enabled expression in a recombinant form from a single plasmid. All ten antibodies gave specific staining patterns within formalin-treated embryonic zebrafish brains, demonstrating that this generalised approach is particularly efficient to elicit antibodies that stain native antigen in fixed wholemount tissue. Finally, we show that additional tags can be easily added to the recombinant antibodies for convenient multiplex staining. The antibodies and the approaches described here will help to address the lack of well-defined antibody reagents in zebrafish research. Copyright © 2014 The Authors. Published by Elsevier Inc. All rights reserved.

Original language	English
Pages (from-to)	527-533
Number of pages	7
Journal	BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS
Volume	456
Issue number	1
Early online date	6 Dec 2014
DOIs	https://doi.org/10.1016/j.bbrc.2014.11.123
Publication status	Published - 2 Jan 2015

Keywords

first & last 3 (QQ only)
first & last (all papers)

Access to Document

10.1016/j.bbrc.2014.11.123

Cite this

@article{9b93ea18202f40a695928938a6ee417f,

title = "A panel of recombinant monoclonal antibodies against zebrafish neural receptors and secreted proteins suitable for wholemount immunostaining",

abstract = "Cell surface receptors and secreted proteins play important roles in neural recognition processes, but because their site of action can be a long distance from neuron cell bodies, antibodies that label these proteins are valuable to understand their function. The zebrafish embryo is a popular vertebrate model for neurobiology, but suffers from a paucity of validated antibody reagents. Here, we use the entire ectodomain of neural zebrafish cell surface or secreted proteins expressed in mammalian cells to select monoclonal antibodies to ten different antigens. The antibodies were characterised by Western blotting and the sensitivity of their epitopes to formalin fixation was determined. The rearranged antigen binding regions of the antibodies were amplified and cloned which enabled expression in a recombinant form from a single plasmid. All ten antibodies gave specific staining patterns within formalin-treated embryonic zebrafish brains, demonstrating that this generalised approach is particularly efficient to elicit antibodies that stain native antigen in fixed wholemount tissue. Finally, we show that additional tags can be easily added to the recombinant antibodies for convenient multiplex staining. The antibodies and the approaches described here will help to address the lack of well-defined antibody reagents in zebrafish research. Copyright {\textcopyright} 2014 The Authors. Published by Elsevier Inc. All rights reserved.",

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author = "Nicole Staudt and Nicole M{\"u}ller-Sienerth and Alla Fane-Dremucheva and Yusaf, {Shahnaz P} and David Millrine and Wright, {Gavin J}",

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A panel of recombinant monoclonal antibodies against zebrafish neural receptors and secreted proteins suitable for wholemount immunostaining. / Staudt, Nicole; Müller-Sienerth, Nicole; Fane-Dremucheva, Alla et al.
In: BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS, Vol. 456, No. 1, 02.01.2015, p. 527-533.

Research output: Contribution to journal › Article › peer-review

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